Fullerene derivatives have been proposed as free radical scavengers. See, e.g., U.S. Pat. No. 5,648,243 to Chiang. A number of academic investigations have studied water soluble C60 derivatives as potential free radical anti-oxidant therapeutics. See, e.g., Jensen et al., Bioorganic & Medicinal Chemistry, 4:767-79, 1996; Da Ros et al., Croatica Chemica Acta CCACAA 74:743-55 (2001); and Wilson, in “Perspectives in Fullerene Nanotechnology,” Osawa, ed., (Kluwer Academic Publishers, Dorcrecht, Netherlands, 2000); Syrensky, et al., Kopf Carrier #63, (David Kopf Instruments Tujunga, Calif., September 2006).
Chiang and colleagues used polyhydroxlated fullerenes to treat laboratory animals in models for ischemia/reperfusion injury, HIV infection, and for neuroprotection. Y. L. Lai and L. Y. Chiang, J. Autonomic Pharmacol., 17:229, 1997; Schinazi et al., Proc. Electrochem. Soc., 97:10, 1997; Lai et al., World J. Surg., 24:450, 2000; Jin et al., J. Neuroscience Res., 62:600, 2000; Huang et al., Free Radical Biol. Med., 30:643, 2001.
Water soluble alkylsulfonyl fullerene derivatives have been shown to be effective in models of focal ischemia, and for photodynamic therapy. Chi et al., in “Perspectives of Fullerene Nanotechnology,” pp 165-183, E. Osawa ed., (Kluwer Academic Publisher, Great Britain, 2002). A tris-malonate derivative, in which the malonate groups are oriented in an equatorial (eee) configuration, C3, has been demonstrated to be effective in preventing oxidative stress in cultured neurons. These observations were translated into efficacy in a model for amyotrophic lateral sclerosis. This same compound also prolonged the life span of mice fed C3 daily. Dugan et al., P.N.A.S. 94:9434-39, 1997; Dugan et al., Parkinsonism & Related Disorders 7:243-46, 2001; Quick et al., Neurobiol of Aging (electronic publication 2006).
Incorporation of fullerenes into lipid vesicles has been studied. Bensasson et al. (Journal of Physical Chemistry, 98:3492-3500, 1994) described preparing vesicles incorporating C60 in L-α-phosphatidyl-choline purified from egg yolk (Egg-PC). However, the authors reported that they were not able to incorporate more than 3% by weight C60 in Egg-PC liposomes and the preparation was not uniformly reproducible. Moreover, the preparation was not designed to be stable and would not have been suitable for administration as a pharmaceutical, diagnostic, cosmetic, or excipient composition.
Modification of fullerenes to produce vesicles has been reported. Hirsch et al. (Angewandte Chemie International Edition, 39:1845-1848, 1999) described C60 to which six pairs of alkyl chains and one polar group was attached. See also U.S. Pat. No. 7,070,810. Felder et al., Helv. Chim. Acta, 85: 288-319, 2002, described a series of compounds wherein complex amphiphilic adducts are coupled to C60. These compounds were designed to self assemble into monolayer films and could not readily be incorporated in biological membranes. Thus, these compounds would not be desirable for therapeutic biological applications.
Fullerenes, including many modified fullerenes that have been studied, have a tendency toward aggregation that can render a composition unsuitable for use as a therapeutic. For example, Williams et al., Recueil des Travaux des Pays-Bas, 1:72-6, 1996, incorporated C60 into L-α-phosphatidyl-ethanolamine from E. coli (PE). Their procedure intentionally initiated formation of C60 clusters, which the authors reported as essential for reproducible preparations. The authors further reported that incorporation of C60 in PE was limited to 7% with C60 adducts being limited to 3%. The limited incorporation means that these compositions are not desirable for use in vivo. Furthermore, the presence of clusters renders the compositions undesirable for use for in vivo delivery, because of the significant risk of toxicity and lack of uniformity.
Fullerenes are hydrophobic, and are generally not soluble in water, but also have poor solubility in lipids. See, e.g., Braun et al., Fullerenes, Nanotubes and Carbon Nanostructures, 15:311-314, 2007. Generally, C60 and other fullerenes do not dissolve in lipids or in many common organic solvents (e.g. hexane, or chloroform).
Kato et al., Chem & Biodiv., 2:1232-1241, 2005, describe C60 with bis mannopyranosyl adducts. These structures are amphiphilic, but the hydrophobic moiety is the C60 cage, which is not lipophilic. Hydrophobic forces cause the C60 bis mannopyranosyl adducts to aggregate. Those aggregates are quite different from biological lipid membranes. The molecules did not associate with dipalmitoyl phosphatidyl choline phospholipid bilayers. Similar aggregation of C60 porphyrin adducts has been described by Georgakilas et al, Proc. Nat. Acad. Sci., 99:5075-5080, 2002. Such aggregates are not comparable or compatible with biological membranes.
Use of fullerenes as antioxidants, particularly in a therapeutic context, has not been adopted, primarily because of problems inherent in the fullerenes and problems in preparing a pharmaceutically acceptable composition and delivery system. Thus, there remains a need in the art for improved fullerene based antioxidants and appropriate delivery systems.